For many years, human ?? T cells were thought solely to recognize peptides bound to MHC proteins. Early stages of work on this parent grant provided broad evidence that non-polymorphic CD1 proteins (CD1a, CD1b, CD1c) display mycobacterial lipids to human ?? T cells. After generating working tetramers comprised of human CD1a, CD1b and CD1c proteins, our published data show that polyclonal, lipid-specific T cells exist as cell populations in tuberculosis (TB) patients and possess effector functions (TNF and IFN-?) that are known to protect the human host. Further, CD1c tetramers detected polyclonal ?? T cells recognizing mycobacterial mycoketides. CD1b tetramers detected a previously unknown human T cell type: germline-encoded mycolylreactive (GEM) T cells. GEM T cells have a degree of T cell receptor (TCR) conservation, which is equivalent to NKT cells or MAIT cells, but GEM T cells function to recognize CD1b. Building on these new reagents and recent success in solving the crystal structure of CD1a-lipid-TCR, the renewal will carry out detailed biophysical analyses of ternary TCR-CD1-mycobacterial lipid interactions involving both ?? and ?? TCRs. Turning to translational questions in TB disease, we will collaborate with Socios En Salud to determine the immunodominant mycobacterial lipids stimulating human response in latent and active TB. Over the past 3 years, we validated guinea pig CD1b and CD1c tetramers and anti-CD1 antibodies to create a tractable small animal model of in vivo T cell response to CD1. Now we propose to measure the duration and magnitude of response to lipid antigens and determine whether CD1 presented lipids can protect against M. tuberculosis infection. Finally, based on the discovery of GEM T cells, we propose to detect networks of interdonor conserved TCRs in the humans. These studies move beyond NKT cells and MAIT cells to detect stereotyped human T cell response to CD1a, CD1b and CD1c bound to mycobacterial lipids. Such studies seek to reveal a currently unappreciated structure of a public human T cell response. Such a system would change general views of the structure of the human T cell repertoire and the particular TCRs identified might be used for immunodiagnosis of TB infection.